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Issue 28, 2019
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PEGylation as an efficient tool to enhance cytochrome c thermostability: a kinetic and thermodynamic study

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Abstract

Cytochrome-c from equine heart was kinetically and thermodynamically investigated either in its native (Cyt-c) or PEGylated forms with different PEGylation degrees (Cyt-c–PEG-4 and Cyt-c–PEG-8). Maximum activities were observed at 80 °C, and the irreversible deactivation was well described by first-order kinetics. The results of activity at different temperatures were used to estimate the activation energy of the catalysed Cyt-c reaction (E* = 10.22 ± 0.40, 7.51 ± 0.06 and 8.87 ± 0.29 kJ mol−1 for Cyt-c, Cyt-c–PEG-4 and Cyt-c–PEG-8) and the standard enthalpy variation of enzyme unfolding (Image ID:c9tb00590k-t1.gif = 33.82 ± 4.92, 109.4 ± 13.1 and 58.43 ± 3.11 kJ mol−1 for Cyt-c, Cyt-c–PEG-4 and Cyt-c–PEG-8, respectively). The results of residual activity tests allowed estimating the activation energy (Ed* = 50.51 ± 1.71, 72.63 ± 0.89 and 63.36 ± 1.66 kJ mol−1 for Cyt-c, Cyt-c–PEG-4 and Cyt-c–PEG-8), enthalpy (ΔH), entropy (ΔS) and Gibbs free energy (ΔG) of the enzyme irreversible denaturation. The higher enthalpic contributions of PEGylated conjugates and the increase in ΔG, compared to the native protein, indorsed the protective role of PEGylation. Negative values of ΔS suggested the occurrence of an aggregation phenomenon by increasing the temperature, which was confirmed by circular dichroism. The estimated thermodynamic parameters suggest that PEGylated Cyt-c forms have enhanced thermostability, which would be of great significance for industrial biosensing applications.

Graphical abstract: PEGylation as an efficient tool to enhance cytochrome c thermostability: a kinetic and thermodynamic study

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Publication details

The article was received on 25 Mar 2019, accepted on 17 Jun 2019 and first published on 18 Jun 2019


Article type: Paper
DOI: 10.1039/C9TB00590K
J. Mater. Chem. B, 2019,7, 4432-4439

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    PEGylation as an efficient tool to enhance cytochrome c thermostability: a kinetic and thermodynamic study

    J. H. P. M. Santos, G. Carretero, S. P. M. Ventura, A. Converti and C. O. Rangel-Yagui, J. Mater. Chem. B, 2019, 7, 4432
    DOI: 10.1039/C9TB00590K

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